This project seeks to define the biochemistry and cell biology of endothelial basement membrane degradation by malignant cells. Basement membranes elaborated by cultured endothelial cells of bovine and human origins and labeled with radioactive precursors in specific components will be prepared using novel techniques developed in this laboratory. These membranes may be prepared in large numbers from both arterial and venour endothelial cells and analysed by sequential enzyme hydrolysis, gel electrophoresis, electron microscopy and immunofluorescence. Human fibrosarcoma cells with the ability to degrade the membranes will be grown in direct contact with the substrates and the enzymology and biochemistry of membrane breakdown determined. In particular, we wish to define the interrelationships between the proteases produced by the fibrosarcoma cells (e.g. plasminogen activator and collagenase) and their relevance to connective tissue breakdown. Subsequently, other malignant cells derived from both carcinomas and sarcomas with differing metastatic potentials will be cultured in direct contact with the labeled basement membranes so that the relevance of protease production to tumor invasion may be better defined. Certain normal cells with invasive potentials such as macrophages and endothelial cells will also be examined for their degradative abilities. Additionally, comparisons will be made of the abilities of malignant cells to degrade arterial, venous and capillary membranes and also their abilities to degrade a pericellular matrix produced by cultured human fibroblasts. These studies in which the cells to be tests will be grown in direct contact with the radioactive insoluble proteins previously produced by other cultured cells, coupled with quantitative techniques, offer a new approach for the study of invasion.